miniPCR bio
TM
Chopped! Using CRISPR/Cas9 to cut DNA - Instructor's and Student's Guide
Version: 1.0 - Release: May 2022 - © 2022 by miniPCR bio™
P./9
Instructor's Guide
• Distribute supplies and reagents to lab groups
Check At the start of this experiment, every lab group should have: Amount
Nuclease-Free Water 20 μl
Reaction Buffer
25 μl
gRNA1 10 μl
gRNA2 10 μl
DNA Sample 25 μl
Proteinase K Solution 20 μl
1X Cas9 Nuclease working solution (on ice) 20 μl
Plastic tubes (see note below for tube size) 4
2-20 μl micropipette
1
Micropipette tips
at least 19
37 °C heat source
Aliquot reagents
20 μl
Nuclease
Free-Water
Water
25 μl
Reaction
Buffer
Buffer
10 μl
gRNA1
10 μl
gRNA2
gRNA2
25 μl
DNA Sample
DNA
20 μl
Proteinase
K Solution
ProK
20 μl
1X Cas9 working
solution (on ice)
Cas9
Aliquot reagents
• For each lab group, label and dispense the
following reagents into seven labeled 200 μl
tubes.
- Nuclease-Free Water, 20 μl
- Reaction Buffer, 25 μl
- gRNA1, 10 μl
- gRNA2, 10 μl
- DNA Sample, 25 μl
- Proteinase K Solution, 20 μl
- 1X Cas9 Nuclease working solution,
(5X Cas9 Nuclease concentrate diluted
with 160 μl Nuclease-Free Water as
described above), 20 μl stored on ice
Note: If aliquoting more than one hour in
advance, store aliquots in the freezer until
use.
Note: Reactions must be incubated at 37 °C for 15 minutes. Students may use either 0.2 ml or 1.7 ml plastic tubes for their
reactions, but should use the size that is compatible with your 37 °C heat source.
